1. Field of the Invention
In general, the invention relates to the detection of analytes in samples. More specifically, the invention relates to the detection of analytes using more than one label to detect the analyte over a broad concentration range.
2. Description of Related Art
Various analytical procedures and devices are commonly employed in specific binding assays to determine the presence and/or amount of substances of interest or clinical significance which may be present in biological or non-biological fluids. Such substances are commonly termed “analytes” and can include, for example, antibodies, proteins, drugs, hormones, cells, and nucleic acids.
Specific binding assays incorporate specific binding members, typified by antibody and antigen immunoreactants, wherein one member of the specific binding pair is labeled with a signal-producing compound (e.g., an antibody or an antigen labeled with an enzyme, a fluorescent compound, a chemiluminescent compound, a radioactive isotope, a direct visual label, etc.). Typically in a sandwich assay the test sample suspected of containing analyte can be mixed with a labeled anti-analyte antibody, i.e., conjugate, and incubated for a period of time sufficient for the immunoreaction to occur. The reaction mixture is subsequently analyzed to detect either that label which is associated with an analyte/conjugate complex (bound conjugate) or that label which is not complexed with analyte (free conjugate). As a result, the amount of label in one of these species can be correlated to the amount of analyte in the test sample. In a competitive assay the test sample is mixed with either a labeled antigen or a labeled antigen analog and these labeled compounds compete with the analyte in the test sample for binding sites on the antibody. The ratio of labeled compound versus test compound determines the level of signal obtained. High analyte concentrations in the test material will result in low signals and vice versa.
Analytes may be present in samples over a broad concentration range. For example, T4 is produced by the thyroid in mammals. A high concentrations of T4 in the blood stream is a marker for hyperthyroidic conditions, and a low concentrations is a marker for hypothyroidic conditions. The difference in T4 concentration between hyperthyroidic and hypothyroidic conditions can be as great as ten fold. Numerous other analytes may be present in biological samples in broad concentration ranges, for example, drugs of abuse, therapeutic drugs, cortisol, HGH, HCG, LSH, TSH, the TORCH panel antigens and the like. While several types of detection systems are available, no one system has been able to simply and easily measure a broad concentration range of T4 or other analytes that may be present in the blood or other samples without resorting to some form of mathematical manipulation. These manipulations include, for example, the use of multiple standard curves to standardize the entire concentration range, or use of some form of a ‘fudge’ factor to alter the standard curve at different points across the concentration range. Alternatively the assay conditions may be altered so that differently optimized assays are run at different points through the concentration range.